Sequence specific binding between proteins and nucleic acids plays a critical role in cells, such as in regulation of gene expression. Due to its major role in all biological processes, the nucleic acid-protein interaction is considered an important drug target. Characterization of such nucleic acid-protein interactions have been largely dependent on gel mobility shift assays, DNase I footprinting assays, or filter binding assays. Although these assays can detect the sequence specificity of the binding event, they may also require the use of labels, such as dyes or radioactive labels. The use of labels on the probe or the target not only adds multiple and tedious steps to the synthesis of the sensor and the method of detection before the assay is performed, but may also be hazardous to living organisms in the environment. In addition, they are often not accurate in determining the binding affinity between the protein and the DNA.
Surface Plasmon Resonance (SPR) spectroscopy, which is a label-independent technique, has become an alternative to label-dependent techniques for studying nucleic acid-protein interactions. However, SPR spectroscopy is expensive to conduct, requiring highly specialized equipment and costly consumables in order to carry out the measurements. This makes SPR spectroscopy unsuitable for fast and cost-effective screening of nucleic-acid protein interactions. This is particularly important for applications such as screening of drug molecules where fast, cost-effective and efficient screening of a plurality of drugs is necessary. In addition, SPR is limited by the fact that only certain size molecules can be measured for binding-interactions, restricting its utility for measuring a wide range of possible interactions.
Fluorescence anisotropy (FA) is an alternative method for studying nucleic acid-protein interactions and for screening low molecular weight ligand inhibitors for the protein binding to nucleic acids. However, FA, like many other methods based on the use of an organic dye, requires attachment of a fluorophore to small molecules that act as probes or ligands for the protein. This can become an extensive iterative process, involving numerous tests for reactions that may occur at multiples sites on the probe or ligand molecule. Furthermore, the optimization of linker length and position can be both time- and labour-intensive. In addition, problems such as steric hindrance may arise upon addition of the fluorescent group to the probe or ligand, which may affect its affinity for the protein. Some fluorophores may also suffer from low inherent photostability, self-quenching and low quantum yields, or only show high sensitivity within a certain pH range. These drawbacks can make the assay inefficient and ineffective.
There is therefore a need to provide a sensor for nucleic acid-protein interaction which may at least partially ameliorate one or more of the disadvantages described above.